RNA干扰沉默蛋白激酶B基因表达对移植静脉内膜增生的影响  

The effect of expression of RNA interference targeting PKB mediated by nanoparticles on intimal hyperplasia in vein graft of rats

作　　者：于维东[1] 胡新华[2] 张志深[2] 李铁民[1] 杨军[2] 张强[1] 

机构地区：[1]中国医科大学附属第一医院干部诊疗科,辽宁沈阳110001 [2]中国医科大学附属第一医院外科,辽宁沈阳110001

出　　处：《中国普通外科杂志》2008年第1期57-60,共4页China Journal of General Surgery

基　　金：国家自然科学基金资助项目(30400435,30672048);辽宁省教育厅科学技术研究项目(2004F76)

摘　　要：目的观察纳米粒子包载的靶向蛋白激酶B(PKB)基因的shRNA表达载体局部转染对大鼠移植静脉内膜增生的影响。方法应用聚乳酸聚乙醇酸共聚物(PLGA)和聚乙烯醇(PVA)包载PKB的RNA干扰基因载体,制备纳米级粒子混合物。建立自体颈静脉-颈总动脉移植模型共72只,随机分成转基因组、空载体组和对照组。分别于术后3,7,14,28 d取材;常规HE及Verhoeff染色,用North-ern blot和Western blot检测PKB基因的mRNA及蛋白的变化,HE和Verhoeff染色观察内膜厚度,TUNEL法观察血管平滑肌细胞(VSMC)凋亡的动态变化。结果转基因组内膜中PKB基因的mRNA及蛋白产物表达较其他两组明显减少(P<0.05);术后7,14,28 d转基因组静脉内膜增生厚度较其他组明显减少(P<0.01);转基因组细胞凋亡率较其他组明显增高(P<0.05)。结论纳米粒子可以作为转基因载体;沉默PKB基因表达能有效地抑制自体移植静脉内膜的增生,促进VSMC的凋亡。Objective To study the effect of expression of RNA interference targeting protein kinase B （ PKB ） gene transfection mediated by nanoparticles （NP） on intimal hyperplasia （ IH ） in vein grafts of rats. Methods Nanoparticle PKB short hairpin RNA （shRNA） gene complex was prepared with PLGA and PVA. Autogenous vein graft model was established in 72 rats by transplanting internal jugular vein to carotid artery. The models were randomly divided into 3 groups ： （ 1 ） PKB shRNA group ： PKB-shRNA gene mediated by NP were transfected into the veins before anastomosis. （2） Empty vector group： the veins were transfected by empty vector mediated by NP. （ 3 ） Control group （ no transfection）. The grafted veins were harvested 3, 7, 14 and 28 days after the operation respectively. The mRNA and protein expression of PKB were determined by Northern blot and Western blot. IH was observed by HE and Verhoeff stain. The presence of apoptotic VSMC was detected by TUNEL stain. Results Compared to empty vector group and control group, PKB shRNA group had less expression of mRNA and protein of PKB gene （ P 〈 0.05 ） , less IH in the vein graft especially during 7 - 28 d （ P 〈 0.01 ） , and more apoptofic VSMC （ P 〈 0.05 ）. Conclusions Nanoparticle is an effective gene transfecting carrier, and RNA interference targeting PKB expression can prevent IH and promote VSMC apoptosis after vein grafting.

关 键 词：蛋白激酶B 短发夹环RNA 内膜增生 纳米粒子 

分 类 号：R654[医药卫生—外科学] R337.1[医药卫生—临床医学]